DISCUSSION

The use of sensitive NAATs compared to traditional virus culture methods has reduced the number of specimens required to confirm the presence of a virus in an outbreak investigation, and multiplex NAATs have allowed the simultaneous detection of multiple viral agents. Restricting initial testing to 3 specimens could significantly reduce laboratory costs while maintaining sensitivity for detecting an outbreak agent. Despite testing specimens from different provinces and using different testing platforms and algorithms, >95% of all outbreaks for which an agent was detected had detected an agent within the first 3 specimens. For influenza, an agent was detected among the first 3 specimens in 96% of outbreaks. Although another study deemed 4 specimens optimal, the findings of marginal increases in virus detection when testing >3 specimens by NAAT mirror the findings here.Reference Peci, Marchand-Austin, Winter and Gubbay ²

During the time period studied, testing 3 specimens per outbreak would have reduced the number of specimens tested by 25.5% in Nova Scotia and 37.8% in Alberta. The associated resources could have been redirected to test additional specimens from outbreaks with few submissions. It is possible that in some outbreaks specimens were not collected because the time lag from symptom onset was considered too long. While specimen collection ideally occurs within 48 hours of symptom onset, this guideline can likely be extended, as both culturable virus and viral RNA can be detected for longer periods of time. ^{1 ,} Reference Carrat, Vergu and Ferguson ^{7 ,} Reference Suess, Remschmidt and Schink ⁸

These findings have several limitations. Like others, ¹ this study ranked specimens by accession time in the laboratory information system. The fact that 2 laboratories within 2 distinct provinces had such strikingly similar findings supports the validity of the results and the unlikelihood that these are chance findings. Second, clinical information was not available, limiting our ability to compare characteristics by viruses detected. Third, classification of outbreak specimens was dependent on proper identification and recording. Similar to other data,Reference Peci, Marchand-Austin, Winter and Gubbay ² we found mixed pathogens in a considerable proportion of outbreaks. However, 3 specimens may not be suitable when mixed pathogens are identified. These situations may require further testing to inform infection control and public health practices, where simultaneous outbreaks or pseudo-outbreaks may be present. Also, further research is required to understand the clinical and epidemiological relevance of respiratory virus coinfection.

Optimal specimen number ultimately depends on sensitivities and specificities of assays in place as well as testing algorithms. However, as an initial step, the routine testing of 3 specimens in an outbreak allows for sensitive detection of influenza and an understanding of the other viruses that cause respiratory outbreaks, all while containing costs. In addition, testing high numbers of specimens can potentially lead to false-positive identification of an outbreak etiologic agent, further supporting the guidance to limit testing numbers.Reference Duizer, Peilaat, Vennema, Kroneman and Koopmans ^{9 ,} Reference Fisman, Greer, Brouhanski and Drews ¹⁰

In summary, these findings demonstrate that 3 specimens is the ideal number when using NAAT to initially detect viral respiratory pathogens in an outbreak investigation. The focus of this study was to determine the minimum number of specimens required to identify a viral pathogen, and these results do not rule out the need for further testing in some circumstances, such as persistence of an outbreak over a long time period; no agent detected; detection of multiple pathogens and/or coinfection; concern related to illness severity; or, for influenza, suspicion of antiviral resistance. Further research is also required to understand the epidemiology of noninfluenza respiratory viruses and to understand how the reporting of these viruses influences medical interventions and/or public health measures.